De novo aging-related NADPH diaphorase positive megaloneurites in the sacral spinal cord of aged dogs.

We investigated aging-related changes in nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) in the spinal cord of aged dogs. At all levels of the spinal cord examined, NADPH-d activities were observed in neurons and fibers in the superficial dorsal horn (DH), dorsal gray commissure (DGC) and around the central canal (CC). A significant number of NADPH-d positive macro-diameter fibers, termed megaloneurites, were discovered in the sacral spinal cord (S1-S3) segments of aged dogs. The distribution of megaloneurites was characterized from the dorsal root entry zone (DREZ) into the superficial dorsal horn, along the lateral collateral pathway (LCP) to the region of sacral parasympathetic nucleus (SPN), DGC and around the CC, but not in the cervical, thoracic and lumbar segments. Double staining of NADPH-d histochemistry and immunofluorescence showed that NADPH-d positive megaloneurites co-localized with vasoactive intestinal peptide (VIP) immunoreactivity. We believed that megaloneurites may in part represent visceral afferent projections to the SPN and/or DGC. The NADPH-d megaloneurites in the aged sacral spinal cord indicated some anomalous changes in the neurites, which might account for a disturbance in the aging pathway of the autonomic and sensory nerve in the pelvic visceral organs.

Cerebral Ischemia/Reperfusion Injury and Pharmacologic Preconditioning as a Means to Reduce Stroke-induced Inflammation and Damage.

Stroke is one of the leading causes of death and long-term serious disability. Current therapeutic strategy is limited to thrombolytic agents, consequently, boosting endogenous neuroprotective mechanisms of the brain to protect itself against harmful stimuli and restore from damages are widely studied. Preconditioned brain to tolerate cerebral ischemia/reperfusion injury could be initiated by several different pharmacological and mechanical strategies, such as ischemic preconditioning, ethanol pharmacological preconditioning and other pre- and post-conditions, such as remote ischemic preconditioning and exercise preconditioning. In this article, we will discuss the major mechanism of ischemia/reperfusion injury and provide an overview of preconditioning in all its various forms, describe the underlying mechanisms and review the recent clinical application of this emerging neuroprotective strategy.

Moderate Ethanol-Preconditioning Offers Ischemic Tolerance Against Focal Cerebral Ischemic/Reperfusion: Role of Large Conductance Calcium-Activated Potassium Channel.

The mechanism underlying moderate ethanol (EtOH)-preconditioning (PC) against ischemic brain injury remains unclear. We evaluated the role of large conductance calcium-sensitive potassium (BKCa) channels in EtOH-PC. Almost one hundred and ninety normal adult SD rats (8 to 10 weeks, 320-350 g) were enrolled in this study. Ischemic/reperfusion (I/R) brain injury was induced in rats by middle cerebral artery occlusion for 2 h followed by reperfusion for 24 h. EtOH or the BKCa channel opener, NS11021, was administered 24 h before I/R with or without pre-treatment with the BKCa channel blocker, paxilline. Infarct volumes were measured by tissue staining and imaging, and neurological functions were assessed by a scoring system. The expression of BKCa channel subunit α was detected by Western blotting, and cell apoptosis was assessed using staining. Prior (24 h) administration of ethanol that produced a peak plasma concentration of ~ 45 mg/dl in rats would offer neuroprotection after cerebral I/R. In addition, the expression of BKCa channel α-subunit was significantly increased 24 h after EtOH-PC (n = 10; control: 2.00 ± 0.09, EtOH: 1.00 ± 0.06; P < 0.5). Compared to I/R, EtOH-PC enhanced the expression of BKCa channel α-subunit both in the penumbra (n = 10; 24 h: I/R: 1.25 ± 0.10, EtOH-PC + I/R: 1.99 ± 0.12; P < 0.01; 4 h: I/R: 1.03 ± 0.03, EtOH-PC + I/R: 1.49 ± 0.05; P < 0.001) and infarct core (n = 10; 4 h: I/R: 1.04 ± 0.04, EtOH-PC + I/R: 1.42 ± 0.05; P < 0.001), improved the neurological function (n = 10; I/R: 14.00 (12.75-15.00), EtOH-PC + I/R: 7.00 (4.75-8.25); P < 0.001), attenuated the apoptosis (n = 10; I/R: 26.80 ± 0.69, EtOH-PC + I/R: 8.46 ± 0.31; P < 0.001), and decreased the infarct volume (n = 10; I/R: 244.00 ± 26.24, EtOH-PC + I/R: 70.09 ± 14.69; P < 0.001) after experimental cerebral I/R. These changes were reversed by paxilline administration. The moderate EtOH-PC protects against I/R-induced brain damage dependent on the upregulation BKCa channels.

Mitochondrial BKCa Mediates the Protective Effect of Low-Dose Ethanol Preconditioning on Oxygen-Glucose Deprivation and Reperfusion-Induced Neuronal Apoptosis.

Ischemia-reperfusion (I/R) injury contributes to the morbidity and mortality of ischemic strokes. As an in vitro model, oxygen-glucose deprivation and reperfusion (OGD/R) exposure induces neuronal injury. Low-dose ethanol preconditioning (EtOH-PC) was reported to alleviate neuronal apoptosis during OGD/R. However, whether the mitochondrial BKCa (mitoBKCa) channel is involved in the neuroprotective effect of EtOH-PC during OGD/R is not clearly defined. This study attempts to explore the mediation of the mitoBKCa channel in the neuroprotective effect of EtOH-PC on OGD/R-induced neuronal apoptosis and the underlying mechanisms. OGD/R model was established using primary cortical neurons that were preincubated with ethanol. Subsequently, the cell viability was measured by CCK-8 assay, and the apoptotic cells were determined by TUNEL assay. Annexin V/7-AAD staining and mitochondrial membrane potential using JC-10 were detected by flow cytometry. Western blot analysis was performed to check the apoptosis-related proteins. In the mixed primary culture, 95% neurofilament-positive cells were cortical neurons. Low-dose EtOH-PC (10 mmol/L) for 24 h significantly attenuated the OGD2h/R24h-induced neuronal apoptosis through activating the BKCa channel. Further investigations suggested that ethanol pretreatment increased the mitochondrial membrane potential (MMP) and downregulated the production of cleaved caspase 3 in OGD/R-injured neurons by activating the mitoBKCa channel. Low-dose ethanol pretreatment significantly attenuated the OGD/R-induced neuronal apoptosis mediated by the mitoBKCa channel which modulated the mitochondrial function by impeding the uncontrolled opening of mitochondrial permeability transition pore (MPTP).

Neuroprotective Effects of Deproteinized Calf Serum in Ischemic Stroke.

Deproteinized calf serum (DCS) may have neuroprotective effects after ischemic stroke. The aim of this study is to investigate whether and how the DCS inhibits neuronal injury following cerebral ischemia. Rats were subjected to 2 h transient middle cerebral artery occlusion (MCAO). One dose of 0.125 mg/gbw DCS was given immediately after reperfusion. Neurological deficit and infarct volume at 24 h post-MCAO in DCS-treated rats were lower than those in vehicle-treated rats (p < 0.0005). In cultured neurons model, cell viability was decreased, and apoptosis was increased by oxygen-glucose deprivation/reperfusion (OGD/R) (p < 0.0005). These effects of OGD/R were attenuated by 0.4 µg/µl DCS (p < 0.05) that were validated by CCK8 cell viability assay, phycoerythrin-Annexin V Apoptosis Detection assay, and TUNEL assay. Furthermore, the increase of intracellular ROS level in cultured neurons was suppressed by DCS (p < 0.05). Compared with cells subjected to OGD/R, the expression level of Bax protein decreased, and bcl-2 protein increased after DSC treatment (p < 0.05). Overall, the neuroprotective effects of DCS following cerebral ischemia may in part be due to decreased ROS production and inhibition of apoptosis.

Synthesis and Evaluations of Novel Apocynin Derivatives as Anti-Glioma Agents.

Apocynin (4-hydroxy-3-methoxyacetophenone) is a natural polyphenolic compound with multiple biological activities. In the present study, a series of apocynin derivatives were designed and synthesized. The in silico ADMET prediction, blood-brain barrier (BBB) penetration assay, anti-NADPH oxidase activity, reactive oxygen species (ROS) levels, and anti-glioma effects of these apocynin derivatives were evaluated. The anti-glioma mechanisms of candidate compounds were studied by flow cytometer and Western blot. The results showed that D31 exhibited higher BBB penetration, increased ROS generations and significant anti-glioma effects both in vitro and in vivo. Further studies showed that D31 inhibited the activations of NF-κB pathway. Overall, our data demonstrated that D31 inhibited growth and induced apoptosis of glioma, which might be caused by ROS-related NF-κB activation. The current study suggested that D31 could be further explored for its potential use in anti-glioma therapy.

C-Fos mapping and EEG characteristics of multiple mice brain regions in pentylenetetrazol-induced seizure mice model.

Purpose: To confirm different local brain activities characterized in pentylenetetrazol (PTZ)-induced seizure model. Methods: we induced seizure response by a single dose of PTZ injection (45 mg/kg, i.p.). Local activity was recorded in different brain regions by EEG in time and c-Fos staining at different time points (0.5 h, 1 h, 2 h, 4 h) after PTZ treatment. Results: EEG recordings showed distinctive features of activation in different brain areas. With the aggravation of behavioral manifestations of seizures, the frequency and amplitude of the discharges on EEG were increasing gradually. The epileptic response on EEG immediately ended after reaching the maximum stage of seizures, followed by a short period of suppression. The labeling of c-Fos was enhanced in the medial prefrontal cortex, the piriform cortex, the amygdala, hippocampal CA1, CA3 and dentate gyrus, but inapparent in the striatum. The most potent changes in c-Fos were observed in cortex, amygdala nuclei, and dentate gyrus. EEG and c-Fos immunolabeling in neuronal activation showed discrepancies in the striatum. For each brain region, the maximum c-Fos labeling was observed at 2 h after injection and diminished at 4 h. The level of c-Fos immunoreactivity was even lower than the control group, which was accompanied by increased labeling of parvalbumin neurons (PVNs). Conclusions: These findings validated PTZ-induced seizure as a seizure model with a specific spatial-temporal profile. Neuronal activity was enhanced and then subsequently inhibited during seizure evolution. Abbreviations: AEDs: anti-epileptic drugs; AF: Alexa Fluor; CA1: Cornu Ammonis area 1; CA3: Cornu Ammonis area 3; DAB, 3: 3P-diaminobenzidine; DAPI: 4',6-diamidino-2-phenylindole; DG: dentate gyrus; EEG: electroencephalogram; GABA: gamma-aminobutyric acid; IEG: immediate early gene; mPFC: medial prefrontal cortex; NAc: nucleus accumbens; PB: phosphate buffer; PBS: phosphate buffered saline; PBST: phosphate buffered saline with Tween; PFA, paraformaldehyde; PTZ: pentylenetetrazol; PVN: parvalbumin neuron; ROI: regions of interest; SE: status epilepticus.

Ketone Bodies Inhibit the Opening of Acid-Sensing Ion Channels (ASICs) in Rat Hippocampal Excitatory Neurons in vitro.

Objectives: Despite the long-term efficacy of antiepileptic drug treatments, frequent attacks of drug-resistant epilepsy necessitate the development of new antiepileptic drug therapy targets. The ketogenic diet is a high-fat, low-carbohydrate diet that has been shown to be effective in treating drug-resistant epilepsy, although the mechanism is yet unclear. In the ketogenic diet, excess fat is metabolized into ketone bodies (including acetoacetic acid, ß-hydroxybutyric acid, and acetone). The present study explored the effect of ketone bodies on acid-sensing ion channels and provided a theoretical basis for the study of new targets of antiepileptic drugs based on "ketone body-acid sensing ion channels." Methods: In this study, rat primary cultured hippocampal neurons were used. The effects of acetoacetic acid, ß-hydroxybutyric acid, and acetone on the open state of acid-sensing ion channels of hippocampal neurons were investigated by the patch-clamp technique. Results: At pH 6.0, the addition of acetoacetic acid, ß-hydroxybutyric acid, and acetone in the extracellular solution markedly weakened the currents of acid-sensing ion channels. The three ketone bodies significantly inhibited the opening of the acid-sensing ion channels on the surface of the hippocampal neurons, and 92, 47, and 77%, respectively. Conclusions: Ketone bodies significantly inhibit the opening of acid-sensing ion channels. However, a new target for antiepileptic drugs on acid-sensing ion channels is yet to be investigated.

Pathogenesis of epileptic seizures and epilepsy after stroke.

Stroke is an important cause of seizures and epilepsy in adults, particularly among the elderly. The incidence of stroke increases yearly as life expectancy increases and the number of patients with post-stroke seizures and epilepsy is also rising. Post-stroke epilepsy accounts for nearly 50% of newly diagnosed epilepsy among patients over 60 years of age. With increasing stroke awareness and advanced treatments, increased attention is paid to post-stroke seizures and epilepsy including its diagnosis and treatment. There has been a plethora of research on the pathogenesis of seizures and epilepsy after stroke. And thus, the research advances in the pathogenesis and related therapeutic targets of post-stroke seizures and epilepsy will be reviewed in this article.

Role of catalpol in ameliorating the pathogenesis of experimental autoimmune encephalomyelitis by increasing the level of noradrenaline in the locus coeruleus.

The endogenous neurotransmitter, noradrenaline, exerts anti-inflammatory and neuroprotective effects in vivo and in vitro. Reduced noradrenaline levels results in increased inflammation and neuronal damage. The primary source of noradrenaline in the central nervous system is tyrosine hydroxylase (TH)­positive neurons, located in the locus coeruleus (LC). TH is the rate­limiting enzyme for noradrenaline synthesis; therefore, regulation of TH protein expression and intrinsic enzyme activity represents the central means for controlling the synthesis of noradrenaline. Catalpol is an iridoid glycoside purified from Rehmannia glutinosa Libosch, which exerts a neuroprotective effect in multiple sclerosis (MS). The present study used an experimental mouse model of autoimmune encephalomyelitis to verify the neuroprotective effects of catalpol. Significant improvements in the clinical scores were observed in catalpol­treated mice. Furthermore, catalpol increased TH expression and increased noradrenaline levels in the spinal cord. In primary cultures, catalpol exerted a neuroprotective effect in rat LC neurons by increasing the noradrenaline output. These results suggested that drugs targeting LC survival and function, including catalpol, may be able to benefit patients with MS.

Identification of two immortalized cell lines, ECV304 and bEnd3, for in vitro permeability studies of blood-brain barrier.

To identify suitable cell lines for a mimetic system of in vivo blood-brain barrier (BBB) for drug permeability assessment, we characterized two immortalized cell lines, ECV304 and bEnd3 in the respect of the tightness, tight junction proteins, P-glycoprotein (P-gp) function and discriminative brain penetration. The ECV304 monoculture achieved higher transendothelial electrical resistance (TEER) and lower permeability to Lucifer yellow than bEnd3. However, co-culture with rat glioma C6 cells impaired the integrity of ECV304 and bEnd3 cell layers perhaps due to the heterogeneity among C6 cells in inducing BBB characteristics. The immunostaining of ZO-1 delivered distinct bands along cell borders on both cell lines while those of occludin and claudin-5 were diffused and weak. P-gp functionality was only proved in bEnd3 by Rhodamine 123 (R123) uptake assay. A permeability test of reference compounds displayed a similar rank order (digoxin < R123 < quinidine, verapamil < propranolol) in ECV304 and bEnd3 cells. In comparison with bEnd3, ECV304 developed tighter barrier for the passage of reference compounds and higher discrimination between transcellular and paracellular transport. However, the monoculture models of ECV304 and bEnd3 fail to achieve the sufficient tightness of in vitro BBB permeability models with high TEER and evident immunostaining of tight junction proteins. Further strategies to enhance the paracellular tightness of both cell lines to mimic in vivo BBB tight barrier deserve to be conducted.

Low-Dose Ethanol Preconditioning Protects Against Oxygen-Glucose Deprivation/Reoxygenation-Induced Neuronal Injury By Activating Large Conductance, Ca2+-Activated K+ Channels In Vitro.

Increasing evidence suggests that low to moderate ethanol ingestion protects against the deleterious effects of subsequent ischemia/reperfusion; however, the underlying mechanism has not been elucidated. In the present study, we showed that expression of the neuronal large-conductance, Ca2+-activated K+ channel (BKCa) α-subunit was upregulated in cultured neurons exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) compared with controls. Preconditioning with low-dose ethanol (10 mmol/L) increased cell survival rate in neurons subjected to OGD/R, attenuated the OGD/R-induced elevation of cytosolic Ca2+ levels, and reduced the number of apoptotic neurons. Western blots revealed that ethanol preconditioning upregulated expression of the anti-apoptotic protein Bcl-2 and downregulated the pro-apoptotic protein Bax. The protective effect of ethanol preconditioning was antagonized by a BKCa channel inhibitor, paxilline. Inside-out patches in primary neurons also demonstrated the direct activation of the BKCa channel by 10 mmol/L ethanol. The above results indicated that low-dose ethanol preconditioning exerts its neuroprotective effects by attenuating the elevation of cytosolic Ca2+ and preventing neuronal apoptosis, and this is mediated by BKCa channel activation.

Reactivated astrocytes as a possible source of oligodendrocyte precursors for remyelination in remitting phase of experimental autoimmune encephalomyelitis rats.

Multiple sclerosis (MS) is ademyelinating disease in the central nervous system (CNS). Majority of the MS patients show relapsing-remitting disease course. Evidences show that oligodendrocyte precursor cells (OPCs), which remain relatively quiescent in normal adult CNS, play a key role in the remitting phase by proliferation and remyelination. In the present study, we found that spinal cord astrocytesco-expressed progenitor cell marker and oligodendroglial lineage markers in the remittance phase in adult rat experimental autoimmune encephalomyelitis (EAE) model. We suggest that activated astrocyte could de-differentiate into OPCs and re-differentiate into mature oligodendrocytes, raising the possibility that astrocytes can be a potential source of OPCs in the adult demyelinated spinal cord.

Resveratrol sensitizes glioblastoma-initiating cells to temozolomide by inducing cell apoptosis and promoting differentiation.

Glioblastoma-initiating cells play crucial roles in the origin, growth, and recurrence of glioblastoma multiforme. The elimination of glioblastoma-initiating cells is believed to be a key strategy for achieving long-term survival of glioblastoma patients due to the highly resistant property of glioblastoma-initiating cells to temozolomide. Resveratrol, a naturally occurring polyphenol, has been widely studied as a promising candidate for cancer prevention and treatment. Whether resveratrol could enhance the sensitivity of glioblastoma-initiating cells to temozolomide therapy has not yet been reported. Here, using patient-derived glioblastoma-initiating cell lines, we found that resveratrol sensitized glioblastoma-initiating cells to temozolomide both in vitro and in vivo. Furthermore, we showed that resveratrol enhanced glioblastoma-initiating cells to temozolomide-induced apoptosis through DNA double-stranded breaks/pATM/pATR/p53 pathway activation, and promoted glioblastoma-initiating cell differentiation involving p-STAT3 inactivation. Our results propose that temozolomide and resveratrol combination strategy may be effective in the management of glioblastoma patients, particularly for those patients who have been present with a high abundance of glioblastoma-initiating cells in their tumors and show slight responsiveness to temozolomide.

Resveratrol Inhibits the Invasion of Glioblastoma-Initiating Cells via Down-Regulation of the PI3K/Akt/NF-κB Signaling Pathway.

Invasion and metastasis of glioblastoma-initiating cells (GICs) are thought to be responsible for the progression and recurrence of glioblastoma multiforme (GBM). A safe drug that can be applied during the rest period of temozolomide (TMZ) maintenance cycles would greatly improve the prognosis of GBM patients by inhibiting GIC invasion. Resveratrol (RES) is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines. The current study aimed to evaluate whether RES can inhibit GIC invasion in vitro and in vivo. GICs were identified using CD133 and Nestin immunofluorescence staining and tumorigenesis in non-obese diabetic severe combined immunodeficient (NOD/SCID) mice. Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo. The activity of matrix metalloproteinases (MMPs) was measured by the gelatin zymography assay. Western blotting analysis and immunofluorescence staining were used to determine the expression of signaling effectors in GICs. We demonstrated that RES suppressed the adhesion, invasion and migration of GICs in vitro and in vivo. Moreover, we proved that RES inhibited the invasion of GICs via the inhibition of PI3K/Akt/NF-κB signal transduction and the subsequent suppression of MMP-2 expression.

Catalpol induces oligodendrocyte precursor cell-mediated remyelination in vitro.

In demyelinating diseases such as multiple sclerosis, one of the treatment strategies includes remyelination using oligodendrocyte precursor cells (OPC). Catalpol, the extract of radix rehmanniae, is neuroprotective. Using an OPC culture model, we showed that 10 µM catalpol promotes OPC proliferation, cell migration and differentiation into mature oligodendrocytes. The 10 µM catalpol displayed stronger effects on OPCs migration and oligodendrocyte differentiation. These results suggest that catalpol has a potential role in promoting remyelination in demyelinating diseases, and is of therapeutic interest.

Down-regulation of apurinic/apyrimidinic endonuclease 1 (APE1) in spinal motor neurones under oxidative stress.

AIM: Apurinic/apyrimidinic endonuclease 1 (APE1) is an intermediate enzyme in base excision repair which is important for removing damaged nucleotides under normal and pathological conditions. Accumulation of damaged bases causes genome instability and jeopardizes cell survival. Our study is to examine APE1 regulation under oxidative stress in spinal motor neurones which are vulnerable to oxidative insult. METHODS: We challenged the motor neurone-like cell line NSC-34 with hydrogen peroxide and delineated APE1 function by applying various inhibitors. We also examined the expression of APE1 in spinal motor neurones after spinal root avulsion in adult rats. RESULTS: We showed that hydrogen peroxide induced APE1 down-regulation and cell death in a differentiated motor neurone-like cell line. Inhibiting the two functional domains of APE1, namely, DNA repair and redox domains potentiated hydrogen peroxide induced cell death. We further showed that p53 phosphorylation early after hydrogen peroxide treatment might contribute to the down-regulation of APE1. Our in vivo results similarly showed that APE1 was down-regulated after root avulsion injury in spinal motor neurones. Delay of motor neurone death suggested that APE1 might not cause immediate cell death but render motor neurones vulnerable to further oxidative insults. CONCLUSION: We conclude that spinal motor neurones down-regulate APE1 upon oxidative stress. This property renders motor neurones susceptible to continuous challenge of oxidative stress in pathological conditions.

Neural progenitor cell apoptosis and differentiation were affected by activated microglia in spinal cord slice culture.

Neural progenitor cell (NPC) transplantation offers great potential to treat spinal cord injury (SCI). NPCs may replace lost neurons or oligodendrocytes and act as a source of neurotrophic factors to support survival of remaining cells. However, their efficiency was limited by poor survival after transplantation, and they tended more to differentiate into astrocytes, but not neurons and oligodendrocytes. This study investigated whether activated microglia is a factor that contributes to this phenomenon. Organotypic spinal cord slice (SCS) culture was used to mimic the local environment after SCI, and NPCs were co-cultured with them to share the culture medium. After specific depletion of microglia in the SCSs with clodronate loaded liposome, the apoptotic rate of NPCs decreased, more NPCs differentiated into neurons, and glial differentiation was impaired. This suggested that microglia may impair NPC survival, and neuronal differentiation, but improve astrocyte differentiation. In NPC transplantation strategy for SCI, microglia would be manipulated to improve the survival and neuronal differentiation of NPCs.

Improvement for identification of heterophile antibody interference and AFP hook effect in immunoassays with multiplex suspension bead array system.

BACKGROUND: A variety of immunoassays including multiplex suspension bead array have been developed for tumor marker detections; however, these assays could be compromised in their sensitivity and specificity by well-known heterophile antibody interference and hook effect. METHODS: Using Luminex® multiplex suspension bead arrays, we modified protocols with two newly-developed solutions that can identify heterophile antibody interference and AFP hook effect. Effectiveness of the two solutions was assessed in serum samples from patients. RESULTS: Concentrations of 9 tumor markers in heterophile antibody positive samples assayed with Solution A, containing murine monoclonal antibodies and mouse serum, were significantly reduced when compared with those false high signals assayed without Solution A (all p<0.01). With incorporation of Solution H (fluorescent beads linked with AFP antigen), a new strategy for identification of AFP hook effect was established, and with this strategy AFP hook effect was identified effectively in serum samples with very high levels of AFP. CONCLUSIONS: Two proprietary solutions improve the identification of heterophile antibody interference and AFP hook effect. With these solutions, multiplex suspension bead arrays provide more reliable testing results in tumor marker detection where complex clinical serum samples are used.

Minocycline inhibited the pro-apoptotic effect of microglia on neural progenitor cells and protected their neuronal differentiation in vitro.

Neural progenitor cell (NPC) transplantation offers great potential to treat spinal cord injury (SCI), but their efficiency is limited by poor survival and neuronal differentiation after transplantation. In the injury site, microglia may become activated and participate in the inflammation reaction. In vitro studies indicated that activated microglia might impair NPC survival and neuronal differentiation, but resting microglia did not. This study investigated the potential of minocycline to modify the negative effects of activated microglia on NPCs in vitro. First, the direct effects of minocycline on NPCs were tested. The results showed that at the concentration of 10µg/ml or lower, minocycline did not affect NPC survival and proliferation, but impaired neuronal differentiation. Then microglia were activated with lipopolysaccharide (LPS) or treated with LPS plus minocycline (LPSMC), and the effects of conditioned media on NPC apoptosis and differentiation were studied. The results showed that, compared with LPS treatment group, the microglia conditioned media of LPSMC treatment group resulted in a significantly lower apoptotic rate of NPCs, and increased the neuronal differentiation of NPCs. This suggested that minocycline might inhibit the negative effects of microglia on NPCs, and have the potential to support the survival and neuronal differentiation of transplanted NPCs for SCI.